The spirochete Borrelia burgdorferi, the causative agent of Lyme disease, is transmitted to humans and other warm blooded animals through the bite of infected Ixodes spp. ticks. The establishment of B. burgdorferi infection involves numerous interactions between the bacteria and a variety of vertebrate host and arthropod vector tissues. Different bacterial proteins are required at specific points of this infectious cycle, and precise regulation of the synthesis of such proteins is essential for successful infection to occur. We have discovered that these spirochetes utilize a regulatory mechanism to control protein expression patterns that involves a chemical signal known as autoinducer-2 (AI-2). This molecule is produced by the B. burgdorferi LuxS protein, which we have demonstrated to be a functional enzyme. Our preliminary studies suggest that B. burgdorferi can regulate LuxS synthesis. Addition of AI-2 to cultured B. burgdorferi dramatically alters the expression of more than 50 different proteins, increasing expression levels of some proteins, while decreasing expression of others. Through this mechanism, a population of Lyme disease spirochetes may synchronize production of proteins needed for infection processes. We hypothesize that B. burgdorferi uses AI-2 as an important signaling molecule to control expression of proteins during the natural infectious cycle of the Lyme disease spirochetes. The proposed studies will characterize the expression of LuxS during bacterial infection, the mechanisms by which B. burgdorferi controls synthesis of AI-2, and functions of AI-2-regulated proteins. Results from these studies will enhance our understanding of the complex interactions between B. burgdorferi and its hosts, as well as identifying potential bacterial targets for improved therapies to prevent and treat Lyme disease.